Stabilized insulinotropic peptides and methods of use
The present invention provides stably crosslinked insulinotropic polypeptides having superior and unexpected benefits in the treatment of conditions involving abnormal glucose homeostasis, e.g., type 2 diabetes and conditions relating to type 2 diabetes. Such benefits include, but are not limited to, extended polypeptide half-life, enhanced alpha-helicity, improved thermal stability and protease resistance, increased functional activity and pharmacologic properties, improved bioavailability when administered by any route, and improved bioavailability and gastrointestinal absorption when delivered orally, as compared to the corresponding unmodified polypeptides. The invention also provides compositions for administering the polypeptides of the invention, as well as methods for preparing and evaluating the polypeptides of the invention.
1. A cross-linked polypeptide comprising an alpha helix and one or more hydrocarbon staples, wherein each hydrocarbon staple stabilizes the alpha helix of the cross-linked polypeptide, and wherein the cross-linked polypeptide comprises gastric inhibitory polypeptide preprotein (GIPP), gastric inhibitory peptide (GIP), glucagon-like peptide-1 precursor (GLP-1P), glucagon-like peptide-2 (GLP-2), liraglutide, taspoglutide, albiglutide or LY2189265.
2. The cross-linked polypeptide of claim 1 , wherein the number of hydrocarbons is between 1 and 10.
3. The cross-linked polypeptide of claim 1 , wherein the hydrocarbon staple stabilizes the N-terminal half of the polypeptide.
4. The cross-linked polypeptide of claim 1 , wherein the hydrocarbon staple stabilizes the C-terminal half of the polypeptide.
5. The cross-linked polypeptide of claim 1 , comprising at least one hydrocarbon staple within the C-terminal half of the polypeptide and at least one hydrocarbon staple within the N-terminal half of the polypeptide.
6. The cross-linked polypeptide of claim 1 , comprising a first hydrocarbon staple located at position (i, i+3), or (i, i+4) or (i, i+7) relative to the residue positions of the alpha helix of the polypeptide.
7. The cross-linked polypeptide of claim 1 , comprising a first hydrocarbon staple located at position (i, i+3), or (i, i+4) or (i, i+7) relative to the residue positions of the alpha helix of the polypeptide, and a second hydrocarbon staple located at position (i, i+3), or (i, i+4) or (i, i+7) relative to the residue positions of the alpha helix of the polypeptide, with the proviso that the first and second hydrocarbon staple are not located at identical positions.
8. The cross-linked polypeptide of claim 1 , wherein the cross-linked polypeptide possesses a half-life that is at least 2-fold greater than the half-life of a non-cross-linked counterpart polypeptide.
9. The cross-linked polypeptide of claim 1 , comprising at least two hydrocarbon staples which are sequentially arranged and in a stitched configuration whereby the end of the first hydrocarbon staple and the beginning of the second hydrocarbon staple originate at a common residue in the polypeptide.
10. The cross-linked polypeptide of claim 1 , wherein the cross-linked polypeptide possesses a resistance to chymotrypsin in vitro that is at least 2-fold greater than the resistance to chymotrypsin in vitro of a non-cross-linked counterpart polypeptide.
11. The cross-linked polypeptide of claim 1 , wherein the cross-linked polypeptide possesses a resistance to pepsin in vitro that is at least 6-fold greater than the resistance to pepsin in vitro of a non-cross-linked counterpart polypeptide.
12. The cross-linked polypeptide of claim 1 , wherein the cross-linked polypeptide possesses a resistance to a serum protease in vivo that is at least 2-fold greater than the resistance to the serum protease in vivo of non-cross-linked counterpart polypeptide.
13. A pharmaceutical composition comprising a cross-linked polypeptide of claim 1 and one or more pharmaceutically acceptable excipients.
14. The cross-linked polypeptide of claim 1 , wherein each hydrocarbon staple covalently couples two non-natural amino acids within the alpha helix of said polypeptide.